Cyclin E, as a positive regulator of the cyclin-dependent kinase, Cdk2, is a key regulator of cell cycle progression. It can also become, when deregulated, a factor in oncogenesis. However, neither the molecular mechanisms whereby cyclin E stimulates cell cycle progression nor those whereby it participates in oncogenesis have been elucidated. This proposal seeks to investigate the function and regulation of cyclin E in both contexts in normal and tumor-derived human breast epithelial cells, the rationale being that information gained is likely to have eventual implications for prevention, diagnosis and therapy of breast cancer. The research plan consists of four specific aims. The first addresses the issue of cyclin E essentiality and function by eliminating cyclin E expression via novel antisense oligonucleotide technology. The second specific aim, also using antisense oligonucleotide technology, investigates the mechanisms whereby breast epithelial cells regulate cyclin E-associated Cdk2 kinase activity to maintain an appropiate proliferative response. The third specific aim directly addresses the mechanism whereby cyclin E deregulation leads to malignant transformation. Tissue culture models are used to investigate the relationship between cyclin E overexpression and genetic instability, whereas a transgenic mouse model is used to directly probe the mechanisms whereby cyclin E deregulation causes mammary carcinogenesis. Finally, the fourth specific aim seeks to investigate whether deregulation of cyclin E proteolysis or maturation play a role in cyclin E accumulation under conditions associated with oncogenesis. Because reagents and technology are already available for carrying out most of these studies, it is anticipated that rapid progress can be achieved on all research avenues proposed.